Acetylation of ANXA1 reduces caspase-3 activation by enhancing the phosphorylation of caspase-9 under OGD/R conditions

Abstract

SIRT2, a Class III HDACs, aggravates cell damage and activates caspase-3 under oxygen-glucose deprivation/reoxygenation and glucose (OGD/R) conditions. In this paper, we demonstrated the adverse effects of SIRT2 on cells after OGD/R attacks, which were mediated by increased interactions between SIRT2 and ANXA1, and explicated the mechanisms by which acetylated ANXA1 affects the activation and cleavage of caspase-3. We found that the acetylation level of ANXA1 was decreased through the its increased interactions with SIRT2 after the OGD/R insult. The lysine 312 residue (K312) was selected as the target site in ANXA1 because it is associated with SIRT2, and its mimic (K312Q) and silent (K312R) mutants were then established through site mutagenesis. Under OGD/R conditions, the acetylation mimic of K312Q ANXA1 accumulated in the cytoplasm, decreasing the activity levels of caspase-3 and the upstream initiator caspase-9, compared with the levels of WT and K312R ANXA1. Furthermore, K312Q ANXA1 intervened in the interactions of caspase-3 to caspase-9 by increasing the phosphorylation levels of caspase-9 and inhibited its cleavage by downregulating PRKAR2B, a regulatory subunit of protein kinase A (PKA). In this process, K312Q ANXA1 was found to be directly associated with PRKAR2B, diminishing its restriction on the catalytic subunit of PKA.In conclusion, acetylated ANXA1 can promote the phosphorylation of caspase-9 to decrease the activation of caspase-3 by enhancing the expression of a kinase upstream of caspase-9 after the OGD/R stimulation.